This invention relates to a porous membrane capable of promoting cell attachment and growth and to a process for making the same. More particularly, this invention relates to a microporous or ultrafiltration membrane containing a cell growth promoting substance and is formed from a porous membrane substrate wherein the substrate has its basic properties changed after treatment in accordance with this invention and to a process for forming such a membrane.
As used herein, the term "transparent" means microscopically transparent. That is, the membrane is transparent to the extent that normal sized cells, e.g., 5 to 30 microns, can be viewed through the membrane structure with a microscope such as at 50 times to 600 times magnification. The transparency can be specifically quantified by measuring optical density with visible light such as with a spectrophotometer.
In many applications of membrane technology, it is desirable to utilize a biocompatible membrane filter which is mechanically strong, is thermally stable, is relatively inert chemically and is insoluble in most organic solvents. Often, it is desirable that the membrane have surface properties which are radically different from and sometimes incompatible with the bulk properties set forth above. In some instances it is desired to form a porous membrane which has low non-specific protein binding and which is capable of supporting cell attachment and growth in order that cell morphology can be easily monitored merely by microscopic examination of the live cells on the membrane. For example, in cell growth technology it is desirable to effect cell growth on and within the pores of a membrane rather than on a flat surface so that three dimensional cell growth rather than two dimensional cell growth can be effected. This is particularly true when growing epithelial cells such as those derived from the lungs, kidneys, or intestine which demonstrate a distinct polarity.
In the intact organism, growing cells are surrounded by an extracellular matrix in a porous environment. Cell culture on a microporous membrane substrate allows the cells to achieve both anatomical and functional differentiation. Over the past few decades, the most widely used cell substrate has been impermeable tissue culture plasticware which limits the expression of differentiated properties. The impermeable plastic prevents the diffusion and transport of both nutrients, waste products and hormones from and to the basolateral cell surface. The addition of extracellular matrix (ECM) components to standard plasticware permits cultured cells to assume increased levels of differentiation but the impermeable nature of plastic and the severely restricted access to both cellular domains still limit the types of experimental investigations that can be pursued. The combination of ECM component coating and a microporous membrane substrate, however, provides a powerful new research tool to investigate in vitro cellular structure and function under conditions which closely mimic the in vivo cellular environment.
Prior to the present invention, porous membranes were used for cell growth and the extracellular matrix (ECM) was applied individually to the membrane by the user. In addition, a non-permanent polytetrafluoroethylene membrane coated with noncrosslinked collagen is available from Costar Corporation under the tradename Transwell--COL. Since the collagen is not crosslinked, the collagen coating is not permanent and it gradually leaches from the polymer surface. In addition, this simple coating makes the PTFE membrane transiently hydrophilic and also greatly reduces the porosity and permeability of the membrane. It would be desirable to provide a substrate in the form of a membrane which is ready to use in that it is biologically active, is permanently bound to the membrane, and permanently hydrophilic without the requirement on the individual user to apply an ECM to the membrane.
Accordingly, it would be highly desirable, for example, to provide a composite membrane having both desirable bulk physical strength and chemical resistance while having desired surface properties different from the bulk properties. Furthermore, it would be desirable to provide such a membrane, which exhibits very low non-specific protein adsorption, low or no autofluoresence and which promotes viable attachment and cell growth.